The present invention relates to methods and systems for generating cell lineage tree of a multiple cell samples and, more particularly, which can be used for assessing clonality of a tumor.
A multicellular organism develops from a single cell, the zygote, through numerous binary cell divisions and cell deaths. Consequently, the lineage relations between the cells of the organism can be represented by a rooted labelled binary tree (FIG. 1a). Such a tree was reconstructed for the 959 somatic cells of Caenorhabditis elegans by direct observation (1). Understanding the cell lineage trees of higher organisms, especially Human, is a fundamental challenge of many branches of biology (2-10) and medicine (11-15). Development of higher organisms is, however, less deterministic than that of C. elegans and therefore the cell lineage trees of individuals of the same species may vary considerably.
Lineage relations among cells have been studied using a variety of clonal assays (2, 3, 6, 8, 10, 16-23). Such assays act by detecting the progeny of a single founder cell, which has been marked by a unique heritable marker. Some assays mark the founder cell by an invasive technique such as injection of a tracer molecule 16, 18 or retroviral infection 10, which may interfere with the normal growth and the biological function of the marked cell population.
Another approach is to detect the progeny of a founder cell that has been affected by a spontaneous mutation or an epigenetic change. Examples include loss or gain of large genomic fragments (19), mitochondrial DNA mutations (20), T cell receptor gene recombinations (21), X-chromosome inactivation (22), and changes in the number of MS repeat units (15, 23). A clonal assay provides limited lineage information because it only determines whether certain cells belong to the sub-tree of the organism cell lineage tree rooted by the clone founder. More subtle lineage relations have been uncovered using quantitative analysis of multiple somatic mutations, enabling the estimation of tumour age and the reconstruction of human tissue lineage trees. In one study (24), tissue samples from breast cancer patients were analyzed for loss of heterozygosity and mutations in mitochondrial DNA, and the result of this analysis was fed into a phylogenetic algorithm, yielding tissue lineage trees. In a different study (25), mismatch repair (26, MMR) deficient colorectal cancer and adenoma tissue samples (which display MS instability) were analyzed for mutations in MS loci. Tissue samples were assigned genotypes from the modes of their MS allele length distributions, and the genotypes were analyzed according to a model of tumour development, yielding reconstructed lineage trees. These studies have tried to exhibit that in certain tissues, lineage trees can be reconstructed from genomic variability.
It should be noted, however, that the above-described lineage analyses were all effected on heterogeneous tissue samples in which lineage relations are undefined and cannot be represented by a binary tree.
There is thus a widely recognised need for and it would be highly advantageous to have methods of identifying cell-lineage tree which are devoid of the above-limitations.